The focus of our investigation is to continue our attempts to obtain chemical information on the couplig mechanism which links the insulin-receptor interaction with membrane transport systems. We have developed 2 dimensional gel electrophoresis systems to greatly improve resolution of membrane proteins and are now ready to attempt to identify membrane sulfhydryls possibly involved in insulin action. This will be accomplished by incubating intact fat cells with labeled NEM according to intricate protocols described herein and analyzing the results on purified plasma membranes. We also plan to attempt to identify the glucose transport protein(s) and the cytochalasin B binding protein(s) which mediates transport inhibition. We have recently found that insulin activates anion transport (SO2 minus over 4) in fat cell plasma membranes when the hormone is added to intact cells and thus propose to characterize the kinetics of anion transport and to identify the anion transport protein(s) in the fat cell plasma membrane. Our interest in identifying the insulin-sensitive hexose and anion transport system proteins is based on our strategy of attempting to discover the nature of the signal mediating insulin action as we learn the chemistry of the responsive transport systems. We also propose to characterize phosphatidylinositol turnover in fat cell membranes since we have found that insulin selectively enhances the incorporation of palmitate and oleate into this phospholipid. We plan to investigate whether this effect may be primary and related to the ability of insulin to modify transport activity.